Expression and immunoreactivity of a human group a rotavirus Vp4 (original) (raw)
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2017
Rotavirus is a non-enveloped viruses containing double-stranded RNA genetic material. The 11 ds-RNA encodes 6 structural and 6 nonstructural proteins. Three major structural capsid proteins of human rotavirus RV4 (VP2, VP6, and VP7) had been isolated, cloned and transfected into Vero cell. VP2 (VP2LRV4), VP6 (VP6LRV4), and VP7 (VP7LRV4) was isolated with the size of coding sequence (CDS) 2673 bp, 1194 bp, and 981 bp, respectively. In silico analysis showed that VP2LRV4, VP6LRV4, and VP7LRV4 protein sequences (prediction) comprised essential subdomain and residues to construct triple layer structural capsid protein. Gene transcription and protein expression of VP2LRV4, VP6LRV4 and VP7LRV4 in Vero cell were examined by RT-PCR and immunofluorescence assay (IF). Transcriptional analysis led to the detection of VP6 and VP7 cDNA on day 1 post transfection while VP2 cDNA was expressed on day 3 post transfection. VP6 and VP7 protein expression marked with fluorescence luminescence in Vero c...
Characterization of VP4 and VP7 of a Murine Rotavirus (YR-1) Isolated in Japan
Japanese Journal of Medical Science and Biology, 1995
Nucleotide and deduced amino acid sequences of the outer capsid proteins VP4 and VP7 of a murine rotavirus strain (YR-1) isolated in Japan were determined. Comparisons of the VP7 amino acid sequence of YR-1 with other murine rotavirus strains (EB, EW, EC, EL and EHP) (1) showed that YR-1 was highly homologous to EB, EW, EC, EL and EHP. Moreover, YR-1 was more closely related to strains representing G3 than to any other G type. Analysis of the VP4 amino acid sequence revealed that YR-1 was highly homologous to EB, EW, EC and EL [tentatively P17 (1)], and more closely related to EHP [tentatively P18 (1)] than to any other P type. Enzyme immunoassay with monoclonal antibodies against G types (KU-4 and BH49 for G1, S2-2G10 and BW36 for G2, YO-IE2 and BC5 for G3; and ST-2G7 and BE18 for G4) and against a P type (YO-1S3, KU-12H and YO-2C2 for P8) showed no reactivity. These results indicate that YR-1 is highly homologous to EB, EW, EC and EL.
Comparative Immunology, Microbiology and Infectious Diseases, 1994
The molecular basis of pathogenesis in vivo for a virulent mouse rotavirus (MRV) and a less virulent bovine rotavirus (BRV) were compared under in vitro and in vivo conditions. Obvious differences in the mobility of several genomic RNA segments were observed in one-dimensional gels. Under in vitro conditions, partial proteolytic peptide mapping identified differences between the two outer capsid proteins of these virus and no difference in inner capsid protein was observed. Since it has been observed by us and others that the gene coding for VP4 protein plays a significant role in determining virulence, the variability observed in the present study between the 84 k proteins (VP4) provided a basis for further investigations in order to locate a potential virulence determinant. A comparison of the carboxypeptidase digests of the MRV-and BRV-VP4 revealed an area of variability between amino acids 307 and 407, which may represent a site of virulence determinant. Under in vivo conditions the virulence of both parenteral BRV and MRV isolates and their corresponding reassortants (with replaced gene 4) were studied in routine and bovine hosts. Like their parents, BRV and MRV isolates, reassortants obtained by replacement ofgene 4 in BRV with MRV gene 4 indicated that the dose of the virus isolate used and the clinical outcome in vivo was determined by gene segment 4. The implications of these findings to elucidate the molecular basis of pathogenesis of rotaviruses are discussed.
VP4 monotype specificities among porcine rotavirus strains of the same VP4 serotype
Journal of virology, 1991
The porcine rotavirus OSU strain was used to produce monoclonal antibodies (MAbs) directed against the outer capsid protein VP4. From two separate fusions, eight MAbs that inhibited hemagglutination activity of the OSU strain were selected. All MAbs immunoprecipitated both the OSU VP4 protein derived from a lysate of infected MA104 cells and the OSU VP4 protein expressed in Sf9 cells by a recombinant baculovirus. By immunoprecipitation of in vitro-translated OSU gene 4 transcripts of different length, the eight MAbs were found to be specific for the VP8 subunit of VP4. All MAbs neutralized the OSU strain but failed to neutralize human, bovine, and simian rotavirus strains. Antiserum to the expressed OSU VP4 protein was used to study the distribution of VP4 antigenicity among porcine rotaviruses. At least two distinct specificities were identified among 14 rotavirus strains that had been previously assigned to four distinct VP7 serotypes. Five groups of monotype specificities of the ...
Archives of virology, 1997
The genes encoding the outer capsid VP4 proteins of four lapine rotavirus strains, three isolated in the US (ALA, C-11 and BAP-2) and one isolated in Japan (R-2) were sequenced, and the predicted amino acid (aa) sequence was compared to all known rotavirus genotypes. A high degree of aa identity (96.8-98.9%) was found among the American lapine strains, while the Japanese rotavirus strain R-2 shared less aa identity (89.5-90.0%) with the American strains. The four lapine rotaviruses shared the closest aa identity (90.6-94.9%) with the P[14] genotype, consisting of viruses isolated from humans in Italy, Finland and Thailand. These results indicate that the VP4 protein of the four lapine strains are genotype P14, and that among lapine strains there are possibly two subtypes, one represented by the American lapine strains and the other by the Japanese R-2 strain.
Journal of Virology, 1993
Simian rotavirus (RRV) and murine rotavirus (EDIM-RW) differ dramatically in the oral inoculum required to cause diarrheal disease in neonatal mouse pups and in their ability to spread and cause disease in uninoculated littermates. A genetic approach was used to explore the molecular basis of these differences. Reassortant viruses were produced in vivo by coinfecting infant mice with RRV and EDIM-RW. Reassortant viruses were isolated by plaque purification of progeny virus obtained from mouse pup intestines on MA104 cells. The plaque-purified reassortants were evaluated for 50% diarrhea dose (DD50) and for the ability to spread and cause diarrhea in uninoculated littermates. The parental RRV strain had a DD50 of 10(5) PFU per animal, while the EDIM-RW parental strain had a DD50 of less than 1 PFU per animal. RRV never spreads from inoculated to uninoculated littermates and causes disease. Twenty-three reassortants were tested. Of great interest were the reassortants D1/5 and C3/2, w...
Sequence analysis of VP4 genes of wild type and culture adapted human rotavirus G1P[8] strains
Asian Pacific Journal of Tropical Medicine, 2011
To conduct a comparative analysis of the VP4 gene sequences of Indian wild type (06361, 0613158, 061060 and 0715880) and cell culture adapted (06361-CA, 0613158-CA, 061060-CA and 0715880-CA) G1P[8] rotavirus strains.Full-length VP4 genes of each of the four wild type G1P[8] rotavirus strains and their cell culture adapted counterparts displaying consistent cytopathic effect were subjected to RT-PCR amplification and nucleotide sequencing.All four cell culture adapted G1P[8] rotavirus strains showed nucleotide and amino acid substitutions in the VP4 gene as compared to their wild type strains. The number of substitutions however, varied from 1-64 and 1-13 respectively. The substitutions were distributed in both VP5* and VP8* subunits of VP4 gene respectively of permeabilization and hemagglutinating activity. The presence of unique amino acid substitutions was identified in two of the four wild type (V377G, S387N in 061060 and I644L in 0715880) and all four cell culture adapted (A46V in 0613158-CA, T60R in 06361-CA, L237V, G389V and Q480H in 061060-CA and S615G and T625P in 0715880-CA) strains for the first time in the VP4 gene of P[8] specificity. Amino acid substitutions generated increase in the hydrophilicity in the cell culture adapted rotavirus strains as compared to their corresponding wild type strains.Amino acid substitutions detected in the VP4 genes of G1P[8] rotavirus strains from this study together with those from other studies highlight occurrence of only strain and/or host specific substitutions during cell culture adaptation. Further evaluation of such substitutions for their role in attenuation, immunogenicity and conformation is needed for the development of newer rotavirus vaccines.
Rotavirus gene structure and function
Microbiological Reviews, 1989
TABLE 1. Classification of group A rotaviruses based on outer capsid protein VP7' VP7 (G) Strain from following species of origin: serotype Human Animal'
Virology, 2000
During a limited epidemiological study, the serotype specificities of several isolates of bovine rotavirus, exhibiting identical electropherotypes, from a single cattle farm near Bangalore, India, could not be determined using a panel of serotyping monoclonal antibodies (MAbs) specific for G serotypes 1-6 and 10. To determine the genotypes of these isolates, the nucleotide sequences of the genes encoding the outer capsid proteins VP4 and VP7 of two representative isolates, Hg18 and Hg23, were determined. The corresponding gene sequences from the two isolates were identical, indicating that these isolates represented a single strain of bovine rotavirus. Comparison of the VP4 nucleotide (nt) and the deduced amino acid (aa) sequences with those of several human and animal rotavirus strains representing all of the currently recognized 20 different VP4 (P) genotypes revealed low nt and aa sequence identities of 61.0 to 74.2% and 57.9 to 78.2% for VP4. The percentages of amino acid homology for the VP8* and VP5* regions of VP4 were 37.7 to 67.9 and 68.1 to 84.2%, respectively. The nt and aa sequences of the VP7 gene were also distinct from those of human and animal strains belonging to the previously established 14 VP7(G) serotypes (65.9 to 75.5% nt and 59.5 to 77.6% aa identities). These findings suggest the classification of the VP4 and VP7 genes of the bovine isolates represented by Hg18 as new P and G genotypes and provide further evidence for the vast genetic/antigenic diversity of group A rotaviruses.